Myelofibrosis (MF) belongs to a group of closely related chronic blood diseases, myeloproliferative disorders (MPD). Most MF patients have no symptoms or present with non- specific constitutional symptoms (weight loss, fatigue), as such an early-stage diagnosis of MF is challenging. MF is clinically suspected when patients present with an enlarged spleen and/or with anemia, thrombocytopenia, or leukopenia/leukocytosis; however, these are neither sensitive nor specific enough to establish a diagnosis of MF. Histological examination is unable to detect early onset bone marrow fibrosis or predict the progression from pre-fibrotic polycythemia vera (PV), essential thrombocythemia (ET) to MF or the likelihood of malignant transformation of MF to leukemia. IsoPlexis is developing a device to monitor individual multi-secreting hematopoietic cells that will not only be able to analyze bone marrow samples but also capture and analyze rare cell count from easily obtainable peripheral blood samples allowing for early diagnosis and better treatment options. IsoPlexis has already validated the device to CD4 and CD8 cells and seeks to demonstrate its critical value to detecting rare circulating hematopoietic cells. The IsoPlexis device can isolate thousands of individual live cells and analyze 20+ secreted proteins to characterize the complex, cell-specific inflammatory response that is correlative to the MF patient survival. The device measures all safety and efficacy functional cytokines per cell and can predict therapeutic responses and potentially enable patient stratification and improved treatment management. Herein, we propose to use the SBIR direct to Phase II grant to demonstrate the validity of our system for use in characterizing, monitoring and predicting the progression of MPD or its treatment. We propose the following specific aims: AIM 1: Develop a fully packaged single- cell protein secretion assay device to meet commercial usability and validity standards, benchmarking sensitivity, specificity, and sample variability for hematopoietic cells from MF patient bone marrow samples collected at MSKCC. AIM 2: Develop algorithmic software and protocols to adeptly compare single-cell cytokine profiles between patients with polycythemia vera (PV), essential thrombocytosis (ET) and MF. AIM 3: Implement enrichment technology, on-chip, that IsoPlexis is currently developing as an all-in-one device to enrich rare cells from circulating blood and detect single cell activity from that subpopulation.